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目的:建立能表达抗汉滩病毒中和性鼠/人嵌合抗体基因的转基因植株,为汉坦病毒基因工程抗体的研究提供实验基础。方法:将构建的含抗汉滩病毒中和性鼠/人嵌合抗体基因的植物抗体表达质粒3G1MH-pCAMB IA2301,用TSS冻融法导入农杆菌GV3101,利用农杆菌介导的抽真空转基因技术转化野生型拟南芥,获得转基因植株。用PCR、Northernblot检测转基因植株。结果:经PCR分析表明,拟南芥的基因组中有抗汉滩病毒中和性鼠/人嵌合抗体基因整合,并成功获得7株T0代转基因拟南芥。植株提取RNA,经Northernblot分析可见约1 500 bp及800 bp处的目的条带,为编码重、轻链的目的基因。结论:成功地构建了含抗汉滩病毒中和性鼠/人嵌合抗体基因的拟南芥植株,为进一步利用植物表达治疗性抗体奠定了基础。
OBJECTIVE: To establish a transgenic plant expressing chimeric antibody against neutralizing mouse / human Hantaan virus and to provide experimental basis for the study of Hantaan virus genetically engineered antibody. METHODS: The constructed plant antibody expression plasmid 3G1MH-pCAMB IA2301 containing neutralizing mouse / human chimeric antibody against Hantaan virus was introduced into Agrobacterium tumefaciens GV3101 by freeze-thaw method using Agrobacterium tumefaciens. Transformed wild-type Arabidopsis to obtain transgenic plants. Transgenic plants were detected by PCR, Northern blot. Results: PCR analysis showed that there was anti-Hantaan virus neutralizing mouse / human chimeric antibody gene integration in Arabidopsis thaliana genome and seven T0 generation transgenic Arabidopsis thaliana were successfully obtained. The RNA extracted from plants showed the target bands of about 1500bp and 800bp by Northern blot, which were the genes encoding heavy and light chains. CONCLUSION: Arabidopsis thaliana plants containing the chimeric antibody against Hantaan and neutralizing mouse / human chimeric antibody were successfully constructed and laid the foundation for the further utilization of plants for the expression of therapeutic antibodies.